Vitamin D3 intake and serum levels are key determinants in establishing risk for human colon cancer, and vitamin D3 has been shown to inhibit the development of AOM induced aberrant crypt foci, and tumors in ApcMin mice. Most studies of mechanism have focused on colon epithelial cell lines in culture, documenting significant effects on proliferation and/or apoptosis, and pathways associated with these cellular responses. However, we recently discovered that in vitro, vitamin D3 disrupts, in a vitamin D receptor-dependent manner, the ability of macrophages to stimulate colon tumor cell growth, and have dissected a specific mechanism and cascade of signaling events disrupted by vitamin D3 in the macrophages by which they signal to the epithelial cells. We will use a novel VDRflox/flox mouse, in which we will target inactivation of the vitamin D receptor to macrophages/monocytic or epithelial cells by introduction of a cfms-cre or villin-cre transgene, respectively. By introducing these constructs into the Apc1638N/+ mouse, we will determine the contribution of the different cell compartments to the effects of vitamin D3 on intestinal homeostasis and prevention of preneoplastic changes and tumor development, and will determine the effects of vitamin D3 in these mice on the specific cross-talk between pathways we have discovered. Establishment that vitamin D3 targets macrophages, as well as epithelial cells, in vivo, and aborts the signals they send to stimulate tumor epithelial cell growth, will establish a new paradigm regarding how vitamin D3 is chemopreventive for colon cancer. PUBLIC HEALTH RELEVANCE: Low levels of vitamin D3 in humans are linked to higher risk for colon (and other) cancers, and elevating vitamin D3 has been shown to be chemoprotective for intestinal tumor development in humans, and in rat and mouse models of the disease. Most studies of how vitamin D influences tumor development have focused on how it affects the growth of epithelial cells of the tumor, the actual malignant cell that invades and metastasizes. However, tumors are made up of many different cell types, and we have recently discovered that other cells in the tumor (macrophages/monocytic cells) can send specific signals to the epithelial cells to cause them to grow more rapidly, and have dissected just which molecules are involved both in the macrophages and the epithelial cells in the "cross-talk". Strikingly, vitamin D3 can inhibit the ability of the macrophages to make and send these signals, thus indicating that macrophages may be a key target in how vitamin D3 inhibits the development and progression of cancer. We have novel mice which have been genetically manipulated so that only the epithelial cells or the macrophages/monocytic cells can respond to vitamin D3. We will use these to prove that macrophages are also key targets in vitamin D3 inhibition of intestinal tumors, and will also determine whether the molecular targets in these animals are the same as those we have discovered when macrophages and epithelial cells are co-cultured together as purified cells.